Transplanted human neurons derived from a teratocarcinoma cell line (NTera‐2) mature, integrate, and survive for over 1 year in the nude mouse brain

Abstract

Retinoic acid (RA) induces a human teratocarcinoma cell line (NTera-2 or NT2) to give rise exclusively to post-mitotic neuron-like (NT2N) cells, but NT2N cells never acquire a fully mature neuronal phenotype in vitro. To determine whether NT2N cells can mature into adult neuron-like cells in vivo, purified NT2N cells were grafted into different regions of the central nervous system (CNS) of adult and neonatal athymic mice, and the grafts were examined immunohistochemically by light, confocal, and electron microscopy using antibodies to a panel of developmentally regulated neuronal polypeptides. NT2N grafts were distinguished from endogenous mouse neurons with antibodies that recognize human or murine specific epitopes in selected neuronal polypeptides. Viable NT2N cells were identified in > 89% of graft recipients (N = 90), and some grafts survived 14 months. Within 3 weeks of implantation, grafted NT2N cells re-extended their processes, and the location of the grafts (e.g., septum versus neocortex) appeared to determine the extent to which processes were elaborated. Within the early post-transplantation period, grafted NT2N cells expressed the same neuronal polypeptides as their in vitro counterparts. However, between 6 weeks and 4-6 months post-implantation, the grafted NT2N cells progressively acquired the molecular phenotype of fully mature in vivo neurons as evidenced by dramatically increased expression of the most highly phosphorylated isoforms of the heavy neurofilament subunit, and the de novo expression of adult CNS tau. Notably, the time course for the extension of processes and the expression of neuronal polypeptides by NT2N grafts was similar in neonatal and adult mice. Although grafted NT2N cells formed synapse-like structures and elaborated dendrites and axons, these axons remained unmyelinated. Finally, none of the transplanted NT2N cells reverted to a neoplastic state. These studies demonstrate that pure populations of grafted human NT2N cells acquire a fully mature neuronal phenotype in vivo, and that these cells integrate and survive for > 1 year post-implantation in the mouse CNS. These human neuron-like cells are an attractive model system for studies of neuronal development, polarity and transplantation.